When organs of the body are formed, they develop in neatly organized arrays. Often, cell types are separated by connective tissue called basement membranes. In skin, for instance, the superficial layer of epidermal cells adheres to the underlying basement membrane. This skin basement membrane acts as a barrier between the epidermal cells on the outside, and the dermal cells underneath. A similar arrangement of cells occurs in the lining of the gut and in the oral cavity.
Basement membranes have been implicated in the growth, attachment, migration, repair and differentiation of their overlying cell populations. Three layers have been defined in basement membranes: a) the lamina lucida, an electronmicroscopically clear region in close approximation to the overlying cells; b) the lamina densa, an electron dense region of 20-300 nm in width; and c) the sublamina densa which contains anchoring fibrils, microfibrillar bundles and collagen fibers.
Many epithelial cells interact with the underlying extracellular matrix, a network of proteins to which cells attach, via a junction called the hemidesmosome (Staehelin, (1974) Structure and Function of Intercellular Junctions, Department of Molecular, Cellular and Developmental Biology, University of Colorado, Boulder, Colo., 191-283). The hemidesmosome, with its anchored structures including intermediate filaments and anchoring fibrils, forms an adhesion complex.
When cultured on tissue culture plastic in vitro, most epithelial cells do not assemble bona fide hemidesmosomes despite the fact that they appear to express all of the necessary plaque and hemidesmosomal components. Several cell lines, including the 804G and NBT-II rat bladder carcinoma cell lines, are capable of assembling hemidesmosomes in vitro under standard culture conditions (Riddelle et al., (1991) J. Cell Biol., 112:159-168; Hieda et al., (1992), J. Cell Biol., 116:1497). It has also been reported that substratum-induced staining by anti-hemidesmosome antibodies is greatly diminished in 804G cell cultures that enter in vitro wound sites (Riddelle et al., (1992) J. Cell Sci., 103:475-490).
As described in Langhofer et al. (1993) J. Cell Sci., 105:753-764) and in copending allowed application Ser. No. 08/324,367 U.S. Pat. No. 5,547,106, the entire contents of which are hereby incorporated by reference, when epithelial cells unable to themselves form hemidesmosomes are plated on the cell matrix deposited by 804G or NBT-II rat bladder carcinoma cells, hemidesmosome formation is induced.
In addition, U.S. Pat. No. 5,422,264, the entire contents of which are hereby incorporated by reference, discloses that a soluble matrix equivalent produced by 804G cells can also induce attachment and hemidesmosome formation in cells contacted with the soluble matrix. The 804G deposited (insoluble) matrix and soluble matrix contain similar protein components as visualized by immunoblotting. These proteins exhibit significant similarity to human merosin, a laminin A isoform, and to Drosophila laminin A. Allowed copending U.S. application Ser. No. 08/152,460 U.S. Pat. No. 5,510,263, the entire contents of which are hereby incorporated by reference, discloses the enhanced growth of pancreatic islet cells cultured on the 804G deposited matrix.
Molecules structurally similar, if not identical to the 804G matrix are also produced by human cell lines; however, these molecules have not been observed to induce hemidesmosome formation in cells plated thereon. Rouselle et al. (J. Cell Biol., 114:567-576, 1991; Burgeson et al., PCT WO92/17498 and PCT WO94/05316) describe a molecule called kalinin which is secreted into the culture medium by human keratinocytes and enhances keratinocyte cell attachment. Carter et al. (Cell, 65:599-610, 1991; PCT WO95/06660) describe an epithelial ligand complex called epiligrin found in the extracellular matrix of human keratinocytes. In addition, a 600 kDa basement membrane glycoprotein (BM600) secreted into the culture medium by human keratinocytes (Verrando et al., Biochim. Biophys. Acta., 942:45-56, 1988; Hsi et al., Placenta 8:209-217, 1987) is structurally similar to 804G matrix. Although kalinin and epiligrin stimulate adhesion of cells to a substrate, they have not been reported to induce formation of hemidesmosomes.
Any medical device, including indwelling catheters and colostomy tubes, which breach the skin for extended periods of time will result in inflammation and/or infection. It would be particularly desirable to coat the surface of these devices with epithelial cells prior to or after insertion into the skin to prevent these undesirable processes. It would also be desirable to coat surgical meshes with epithelial cells for use in skin allografts. In addition, periodontitis, a severe form of gum disease resulting in destruction of gum tissue epithelium and bone erosion, would be amenable to treatment with dental abutment pieces coated with epithelial cells. This would promote reattachment of detached gum tissue to the tooth surface.
The maintenance of tissues and organs ex vivo is also highly desirable. Tissue replacement therapy is well established in the treatment of human disease. Human epidermal cells can already be grown in vitro and used to populate burn sites and chronic skin ulcers. However, many primary cells and tissues are difficult to establish in vitro on normal tissue culture plastic. Although this problem is partially alleviated by the use of extracellular matrix-coated cell supports, this is only a temporary solution.
Thus, there is a need for trans-epithelial appliances capable of stimulating epithelial cell attachment and spreading and for a composition capable of supporting the viability of tissues and organs maintained ex vivo. The present invention satisfies these needs.